The mTORC2 Component Rictor Contributes to Cisplatin Resistance in Human Ovarian Cancer Cells
Figure 2
CDDP-induced rictor downregulation in CDDP-sensitive cells involves caspase-3-mediated cleavage.
OV2008 and A2780s were pretreated with Z-VAD FMK (10 µM) and Z-DEVD FMK (50 µM) for 30 minutes before and during CDDP challenge (0-10 µM; 24 h) and rictor content and apoptosis were assessed. OV2008 cells treated with CDDP alone exhibited an intact rictor (200 kDa) and two cleaved products (160 kDa and 130 kDa). CDDP decreased intact rictor content and 160 kDa protein but markedly increased levels of the 130 kDa band. Treatment of A2780s with CDDP resulted in down-regulation of intact rictor but increased the level of the 160 kDa protein and had no effect on the 130 kDa protein. Pre-treatment of the cells with the pan-caspase inhibitor (Z-VAD) or the specific caspase-3 inhibitor (Z-DEVD) significantly attenuated the CDDP-induced changes in intact and cleaved rictor contents in both sensitive cells, and CDDP-induced apoptosis was significantly but not completely attenuated by the presence of the inhibitors in both chemosensitive cell lines. Rictor content was normalized against GAPDH (loading control). Results are presented as mean ± SEM (n=3 and n=5 in OV2008 and in A2780s, respectively). *p<0.05, **p<0.01, ***p<0.001 (vs respective controls). Hoechst 33258 staining was used for assessment of apoptosis as mentioned in experimental procedures.